具核梭杆菌对结直肠癌细胞增殖、迁移及上皮间质转化作用的影响

Effect of Fusobacterium nucleatum on proliferation, migration and epithelial-to-mesenchymal transition of colorectal cancer cells

摘要:

【目的】评估具核梭杆菌对人结直肠癌细胞HCT116和人正常结肠上皮细胞HCoEpiC的增殖、黏附、凋亡、迁移、侵袭和上皮间质转化的影响。【方法】本研究用不同感染复数(MOI)Fusobacterium nucleatum ATCC 23726感染人结直肠癌细胞HCT116和人正常结肠上皮细胞HCoEpiC，建立感染模型；用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide，MTT]、平板克隆、细胞划痕及侵袭(transwell)实验检测两组细胞的增殖、迁移和侵袭的变化；用流式细胞仪检测两组细胞凋亡情况；通过Western blotting检测两组细胞上皮标记物上皮细胞钙黏蛋白(E-cadherin)、Catenin δ-1蛋白、间充质标记物N-钙粘蛋白(N-cadherin)和波形蛋白(vimentin)表达水平的变化。【结果】F.nucleatum可促进HCT116细胞增殖，诱导HCT116细胞的迁移和侵袭，但不能引起细胞凋亡；可抑制HCoEpiC细胞的增殖、迁移和侵袭，并加速其凋亡；对HCT116和HCoEpiC细胞表现出很强的粘附能力，致细胞分散和拉长，细胞间粘附减少；使HCT116和HCoEpiC细胞上皮标记物E-cadherin与Catenin δ-1的表达量减少，间充质标记物N-cadherin与vimentin的表达量上升，E-cadherin由细胞膜向细胞质转移。【结论】F.nucleatum可诱导结直肠癌细胞和人正常结肠上皮细胞发生上皮间质转化，但抑制人正常结肠细胞的增殖、迁移和侵袭，表现出与结直肠癌细胞相反的作用。

Abstract:

[Objective] This study aims to investigate the effects of Fusobacterium nucleatum on the proliferation, adhesion, apoptosis, migration, invasion, and epithelial-to-mesenchymal transition (EMT) of colorectal cancer cell line HCT116 and human normal colon epithelial cell line HCoEpiC. [Methods] HCT116 and HCoEpiC cells were stimulated with F. nucleatum ATCC 23726 at different multiplicity of infection (MOI) to establish infection models, and then MTT, plate cloning assay, wound healing assay, and transwell assay were employed to measure cell proliferation, migration, and invasion. Flow cytometry was employed to detect cell apoptosis. The expression levels of E-cadherin, catenin δ-1, N-cadherin, and vimentin were determined by Western blotting. [Results] F. nucleatum promoted the proliferation, induced the migration and invasion, and did not induce the apoptosis of HCT116 cells. However, it inhibited the proliferation, migration, and invasion and accelerated the apoptosis of HCoEpiC cells. F. nucleatum exhibited strong adhesion to HCT116 and HCoEpiC cells, resulting in dispersed and elongated cells and reduced intercellular adhesion. This bacterium down-regulated the expression levels of the epithelial markers E-cadherin and catenin δ-1 and up-regulated those of the mesenchymal markers N-cadherin and vimentin in HCT116 and HCoEpiC cells. In addition, it promoted the transition of E-cadherin from the cell membrane to the cytoplasm. [Conclusion] F. nucleatum can adhere to both normal colon epithelial cells and colon cancer cells and induce EMT. However, it inhibited the proliferation, migration, and invasion of human normal colon epithelial cells, showing the effect opposite to that on colon cancer cells.